Mast cells orchestrate the recruitment of inflammatory cells and initiate and perpetuate allergic responses through their ability to release a wide array of inflammatory mediators, including histamine and other preformed mediators, de novo synthesized arachidonic acid metabolites (leukotrienes and prostaglandins), and numerous proinflammatory cytokines and chemokines. All have been shown to play important roles in the pathogenesis of asthma and its exacerbation. Our recent studies have begun to implicate the potent sphingolipid metabolites, sphingosine-1 -phosphate (S1P) and ceramide-1 -phosphate (C1P) and the kinases that produce them, sphingosine kinases (SphK1 and SphK2) and ceramide kinase (CerK), respectively, in regulation of degranulation of mast cells and their secretion of chemokines and cytokines, and eicosanoid synthesis (particularly PGD2 and CysLT). This proposal is aimed at enhancing understanding of the roles of these sphingolipid metabolites and the enzymes that regulate their levels in human mast cell functions in allergic responses and asthma. In Aim 1, we will examine the involvement of S1P receptors, SphKs, and CerK in amplifying and perpetuating allergic responses of human mast cells. Aim 2 is focused on determining how S1P is secreted by human mast cells. In Aim 3, we will determine the effectiveness of novel inhibitors of SphKs and FTY720 analogues that target eicosanoid production, on human mast cell functions. In Aim 4, we will determine the effectiveness of these novel inhibitors in alleviation of airway hyper-responsiveness in murine asthma models. These studies will further our understanding of the critical role of S1P and C1P in orchestrating human mast cell functions and immune reactions, providing the basis for development of therapeutic agents that target the enzymes that regulate their levels and "pave the way" for the development of potent and specific drugs that potentially could be useful for treating asthma in patients.